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Peroxisome proliferator-activated receptor gamma isoforms alter lipogenic gene networks in goat mammary epithelial cells
Lactation is a highly demanding lipid synthesis and transport process that is crucial for the development of newborn mammals. Peroxisome proliferator-activated receptor-γ (PPARG) was reported to promote adipogenesis and lipogenesis in adipose tissue, its role in the lactating mammary gland is less clear. PPARG is present in two isoforms generated by alternative splicing, PPARG1 and PPARG2. Their roles in ruminant lactation mammary gland have been poorly distinguished. To determine which of these isoforms is more closely associated with the regulation of the lipogenic pathways, their distributions were analyzed and key genes in the mammary lipid network were detected by quantitative PCR (qPCR) after overexpression of the two isoforms in goat mammary epithelial cells (GMEC). Various tissues of goats were collected to assay mRNA expression of PPARG isoforms. The adenovirus pAd-PPARG1 and pAd-PPARG2 were generated. The adenovirus (Ad-GFP) was used as a positive control. GMEC at about 80% confluence were transfected with adenovirus at the same MOI and cultured in the DMEM/F-12, at 37°C in 5% CO2. Transfected GMEC were cultured with ROSI or DMSO at 50 μM after 24 h of the initial culture and then harvested at 48 h (24 h later) for RNA extraction. Distribution analysis indicated that expression of PPARG2 was markedly greater in adipose than mammary gland and PPARG1 is the mainly isoform in goat mammary tissue. Both PPARG isoforms could significantly upregulate the mRNA expression of NR1H3, INSIG1, PLIN2, CD36, SCD, AGPAT6, DGAT1 under the treatment with rosiglitazone (ROSI). They had no significant effect on SCAP, PNPLA2 and PLIN3 in absence of ROSI. PPARG1 increased the SREBF1, FASN and ACACA while PPARG2 downregulated these genes expression. In conclusion, the data suggest that both PPARG1 and PPARG2 could largely affect fatty acid metabolism when stimulated. However, de novo lipogenesis in mammary cells appear more closely related to PPARG1 activation, with this nuclear receptor acting through its control of SREBF1 and to some extent NR1H3.
Keywords:
PPARG; lipogenesis; mammary gland epithelial cells; goat